The past few decades heralded an era of high carbohydrate (CHO) diets, some of which are now known to worsen lipid profiles and increase risk of cardiovascular disease (CVD). Both diet composition and meal frequency affect lipid synthesis and kinetics, but the specific mechanisms remain unsettled. We will test the hypothesis that the conversion of CHO to fat (de novo lipogenesis [DNL]) is a major determinant of postprandial triglyceride (TG) and lipoprotein fluxes leading to higher levels of serum TG and dyslipidemia. We propose to perform controlled metabolic studies that use diet composition and meal frequency to modulate postprandial DNL and assess its impact on TG and apolipoprotein (apo) B fluxes, lipid profiles and lipoprotein particle distribution. We will do so by measuring th individual contributions of very low density lipoprotein (VLDL; from the liver) and chylomicrons (chylos, from the intestine) to overall postprandial lipoprotein-TG production and clearance in overweight and obese volunteers, a population that is at risk for CVD. We will compare the effects of 7 days feeding with two isocaloric diets (randomly assigned) that are either high in sucrose or high in fat on postprandial DNL, VLDL-TG and chylo-TG, and apolipoprotein B100 and B48 fluxes using stable isotope tracers and sophisticated mathematical modeling. Using a randomized crossover design in participants assigned to each diet, we will also compare the magnitude of differences between successive 7-day periods of consuming small frequent meals (nibbling) and ingesting 2 daily meals, with each participant serving as his/her own control. Participants will be housed in a metabolic ward and fed constant weight-maintaining diets to assure dietary adherence and control activity. Liver lipid storage will be measured by magnetic resonance (MR) spectroscopy, visceral fat by MR imaging, and lean and fat mass by dual energy X-ray absorptiometry. To further link the dynamic fluxes to atherogenic risk, the composition of VLDL and chylo remnants, lipoprotein particle sizes, small dense low density lipoprotein and high density lipoprotein will be measured using proprietary HPLC methods and gel electrophoresis. We anticipate that these novel studies will allow us to identify, for the firs time, the mechanisms by which meal composition and frequency affect postprandial apoB and TG fluxes and associated lipid profiles. These findings could inform efforts to develop evidence-based interventions and dietary guidance to mitigate CVD risk.